The long term goal of this program is to identify and characterize the in vivo structures of the contractile proteins during various phases of contraction. Previously we have identified that myosin heads (cross-bridges) must first attach to actin in a weakly bound, non-force-generating configuration. We have proposed that the generation of isometric force results from a structural change in the cross-bridges making a transition from the weakly bound configuration to a strongly bound, force generating configuration. The focus of our research has been to investigate both experimentally and by modelling, the molecular structures of the two types of cross-bridge configurations. We have proposed that the cross-bridges weakly bound to actin assume a flexible, (non-stereo-specific) configuration; while those strongly bound assume a rigid (stereo-specific) configuration. In FY 95 we have developed computational models to derive expected diffraction patterns from three different configurations of the cross-bridges: when there is no interaction between actin and myosin; when the binding is non-stereo-specific; and when the binding is stereo-specific. Calculations suggest that the non-stereo-specific binding of myosin heads to actin is characterized by the appearance of a mixture of myosin and actin based layer lines, instead of myosin layer lines being dominating as in the case of no interaction. The stereo-specific binding is characterized by the disappearance of myosin-based layer lines while the actin based layer lines become dominant. In FY95 we have obtained high resolution diffraction patterns, by using synchrotron radiation sources, from relaxed and rigor fibers. Experimental diffraction patterns can now be compared with the calculated ones to derive proper interpretation of the experimental data. The structure of weakly bound cross-bridges in the presence of calcium was investigated, since several models on contraction mechanism suggest that activation by calcium induces an intermediate structural state in the pathway to force generation. Two dimensional diffraction patterns were obtained from single fibers using ATPgS in the presence and in the absence of calcium. Preliminary results showed that apart from the changes related to the activation process, there is no large scale structural changes upon addition of calcium. This finding had direct implications for our understanding of the pathway to force generation.